Aberrant signaling through the epidermal growth factor receptor (EGFR/erbB 1) is frequently observed in epithelial malignancies. However, it is poorly understood which of the manifold effects of EGFR activation are critical to the malignant phenotype. During the previous grant cycle the applicants have assigned a novel role to the EGFR in support of anchorage-independent epithelial cell survival. Among the many functions of the EGFR this effect is considered to be rate-limiting to the development of advanced epithelial malignancies as it is likely to impair the establishment of metastatic lesions. The overall goal of this application is to define molecular mechanisms and pathways by which EGFR activation controls anchorage-independent epithelial cell survival. Preliminary results by the applicants demonstrate that matrix-independent, EGFR-dependent MEK/MAPK signaling is critically required for this process. Further preliminary results suggest that (i) EGFR activation sustains MAPK activation of suspended keratinocytes, at least in part, by suppressing MAPK phosphatases; (ii) posttranslational modifications of apoptosis regulators through MAPK are likely to contribute to keratinocyte survival in this setting; (iii) MAPK-dependent effects on transcription of apoptosis modifiers further enhances the survival phenotype. These preliminary results provide the basis for three specific aims focusing on EGFR-dependent regulation of MAPK activity and molecular consequences of sustained MAPK signaling as they relate to keratinocyte survival. The hypotheses to be tested are (1) EGFR activation is required to suppress MAPK phosphatases and thus enables sustained MAPK signaling in suspension culture; (2) MAPK-dependent activation of Rsks contributes to posttranslational modifications of Bcl-2 family members and thus supports cell survival; (3) EGFR-dependent MAPK activation in suspension culture further supports cell survival by maintaining transcription of anti-apoptotic genes. The proposed studies are relevant to physiological states and pathological conditions in which epithelial cells need to survive in the absence of optimal matrix interaction ranging from wound healing to neoplasia. Furthermore, they will provide valuable information about EGFR-dependent control of apoptosis susceptibility as it relates to targeting the EGFR in epithelial cancers.